We describe the earliest petrosal bone referred to the South American endemic Notoungulata, from the late Paleocene-early Eocene Beds of Itaboraí, which provides a critical basis for assessing their enigmatic origins. As indicated by our phylogenetic analysis, the fossil belongs to a taxon most likely close to the ancestral root of the Notoungulata. We describe the entire external anatomy of this isolated ear bone and also investigate the bony labyrinth of the inner ear through computed tomographic scan reconstruction. Within Notoungulata, the specimen retains a number of features considered plesiomorphic, such as a narrow medial flange on the tympanic surface, the presence of a petromastoid canal, and a rather deep fossa subarcuata. We also present a survey of the petrosal anatomy of early diverging notoungulates compared with other members of Eutheria. Features regarded as plesiomorphic in Notoungulata comprise a low stapedial ratio, a cavity for the trigeminal ganglion, a secondary crus commune, a ramus superior of stapedial artery (although reduced) and arteria diploëtica magna, and the tegmen tympani pierced by a canal. Derived features include notably a laterally located tensor tympani fossa, a bean-shaped promontorium and adjoining flattened medial flange, and a stapedial fossa poorly separated from the postpromontorial tympanic sinus. A number of derived notoungulate characters are interestingly shared with the extant hyrax Procavia. Further investigation of this anatomical region in other eutherians is required in order to fully exploit the phylogenetic potential of our new observations. This is especially needed for other groups of South American endemic ungulates possibly related to Notoungulata.